A Tie2‐driven BAC‐TRAP transgenic line for in vivo endothelial gene profiling

Santhosh, Devi; Huang, Zhen

doi:10.1002/dvg.22923

Cited by 6 publications

(3 citation statements)

References 37 publications

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“…Mouse transgenic approaches have enabled transcriptional profiling directly in vivo via translating ribosome affinity purification (TRAP) (31)(32)(33)(34)(35)(36). This method relies on the inducible expression of an epitope-tagged ribosomal protein in a cell typespecific pattern, thus facilitating isolation of ribosome-associated transcripts, also known as the translatome (37), for particular cellular subsets directly from a complex mixture of cells.…”

mentioning

confidence: 99%

The in vivo endothelial cell translatome is highly heterogeneous across vascular beds

Cleuren

Ent

Jiang

et al. 2019

Proc. Natl. Acad. Sci. U.S.A.

108

110

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Endothelial cells (ECs) are highly specialized across vascular beds. However, given their interspersed anatomic distribution, comprehensive characterization of the molecular basis for this heterogeneity in vivo has been limited. By applying endothelial-specific translating ribosome affinity purification (EC-TRAP) combined with high-throughput RNA sequencing analysis, we identified pan EC-enriched genes and tissue-specific EC transcripts, which include both established markers and genes previously unappreciated for their presence in ECs. In addition, EC-TRAP limits changes in gene expression after EC isolation and in vitro expansion, as well as rapid vascular bed-specific shifts in EC gene expression profiles as a result of the enzymatic tissue dissociation required to generate single-cell suspensions for fluorescence-activated cell sorting or single-cell RNA sequencing analysis. Comparison of our EC-TRAP with published single-cell RNA sequencing data further demonstrates considerably greater sensitivity of EC-TRAP for the detection of low abundant transcripts. Application of EC-TRAP to examine the in vivo host response to lipopolysaccharide (LPS) revealed the induction of gene expression programs associated with a native defense response, with marked differences across vascular beds. Furthermore, comparative analysis of whole-tissue and TRAP-selected mRNAs identified LPS-induced differences that would not have been detected by whole-tissue analysis alone. Together, these data provide a resource for the analysis of EC-specific gene expression programs across heterogeneous vascular beds under both physiologic and pathologic conditions.

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mentioning

confidence: 99%

The in vivo endothelial cell translatome is highly heterogeneous across vascular beds

Cleuren

Ent

Jiang

et al. 2019

Proc. Natl. Acad. Sci. U.S.A.

108

110

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“…TIE2 (TEK), a member of the receptor tyrosine kinase subfamily, is mainly expressed in endothelial cells (ECs) [ 7 ]. TIE2 plays an important role in regulating angiogenesis, vascular remodeling, maturation, and integrity [ 8 – 10 ].…”

Section: Introductionmentioning

confidence: 99%

AKT/FOXO1 axis links cross-talking of endothelial cell and pericyte in TIE2-mutated venous malformations

Huang

et al. 2020

Cell Commun Signal

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Background: Venous malformations (VMs), most of which associated with activating mutations in the endothelial cells (ECs) tyrosine kinase receptor TIE2, are characterized by dilated and immature veins with scarce smooth muscle cells (SMCs) coverage. However, the underlying mechanism of interaction between ECs and SMCs responsible for VMs has not been fully understood. Methods: Here, we screened 5 patients with TIE2-L914F mutation who were diagnosed with VMs by SNP sequencing, and we compared the expression of platelet-derived growth factor beta (PDGFB) and α-SMA in TIE2 mutant veins and normal veins by immunohistochemistry. In vitro, we generated TIE2-L914F-expressing human umbilical vein endothelial cells (HUVECs) and performed BrdU, CCK-8, transwell and tube formation experiments on none-transfected and transfected ECs. Then we investigated the effects of rapamycin (RAPA) on cellular characteristics. Next we established a co-culture system and investigated the role of AKT/FOXO1/PDGFB in regulating cross-talking of mutant ECs and SMCs. Results: VMs with TIE2-L914F mutation showed lower expression of PDGFB and α-SMA than normal veins. TIE2 mutant ECs revealed enhanced cell viability and motility, and decreased tube formation, whereas these phenotypes could be reversed by rapamycin. Mechanically, RAPA ameliorated the physiological function of mutant ECs by inhibiting AKT-mTOR pathway, but also facilitated the nuclear location of FOXO1 and the expression of PDGFB in mutant ECs, and then improved paracrine interactions between ECs and SMCs. Moreover, TIE2 mutant ECs strongly accelerated the transition of SMCs from contractile phenotype to synthetic phenotype, whereas RAPA could prevent the phenotype transition of SMCs. Conclusions: Our data demonstrate a previously unknown mechanistic linkage of AKT-mTOR/FOXO1 pathway between mutant ECs and SMCs in modulating venous dysmorphogenesis, and AKT/FOXO1 axis might be a potential therapeutic target for the recovery of TIE2-mutation causing VMs.

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“…Current therapies are invasive and rarely curative as lesions tend to recur, so it needs to seek new therapies to alleviate clinical symptoms. TIE2 (TEK), a member of the receptor tyrosine kinase subfamily, is mainly expressed in endothelial cells (ECs) [7]. TIE2 plays an important role in regulating angiogenesis, vascular remodeling, maturation, and integrity [8][9][10].…”

Section: Introductionmentioning

confidence: 99%

AKT/FOXO1 axis links cross-talking of endothelial cell and pericyte in TIE2-mutated venous malformations

Jiang

Si²,

Huang

et al. 2020

Preprint

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Background: Venous malformations (VMs), most of which associated with activating mutations in the endothelial cells (ECs) tyrosine kinase receptor TIE2, are characterized by dilated and immature veins with scarce smooth muscle cells (SMCs) coverage. However, the underlying mechanism of interaction between ECs and SMCs responsible for VMs has not been fully understood.Methods: Here, we screened 5 patients with TIE2-L914F mutation who were diagnosed with VMs by SNP sequencing, and we compared the expression of platelet-derived growth factor beta (PDGFB) and α-SMA in TIE2 mutant veins and normal veins by immunohistochemistry. In vitro, we generated TIE2-L914F-expressing human umbilical vein endothelial cells (HUVECs) and performed BrdU, CCK-8, transwell and tube formation experiments on none-transfected and transfected ECs. Then we investigated the effects of rapamycin (RAPA) on cellular characteristics. Next we established a co-culture system and investigated the role of AKT/FOXO1/PDGFB in regulating cross-talking of mutant ECs and SMCs.Results: VMs with TIE2-L914F mutation showed lower expression of PDGFB and α-SMA than normal veins. TIE2 mutant ECs revealed enhanced cell viability and motility, and decreased tube formation, whereas these phenotypes could be reversed by rapamycin. Mechanically, RAPA ameliorated the physiological function of mutant ECs by inhibiting AKT-mTOR pathway, but also facilitated the nuclear location of FOXO1 and the expression of PDGFB in mutant ECs, and then improved paracrine interactions between ECs and SMCs. Moreover, TIE2 mutant ECs strongly accelerated the transition of SMCs from contractile phenotype to synthetic phenotype, whereas RAPA could prevent the phenotype transition of SMCs.Conclusions: Our data demonstrate a previously unknown mechanistic linkage of AKT-mTOR/FOXO1 pathway between mutant ECs and SMCs in modulating venous dysmorphogenesis, and AKT/FOXO1 axis might be a potential therapeutic target for the recovery of TIE2-mutation causing VMs.

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A Tie2‐driven BAC‐TRAP transgenic line for in vivo endothelial gene profiling

Cited by 6 publications

References 37 publications

The in vivo endothelial cell translatome is highly heterogeneous across vascular beds

The in vivo endothelial cell translatome is highly heterogeneous across vascular beds

AKT/FOXO1 axis links cross-talking of endothelial cell and pericyte in TIE2-mutated venous malformations

AKT/FOXO1 axis links cross-talking of endothelial cell and pericyte in TIE2-mutated venous malformations

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